Regulation of swelling-activated Cl^- channels by P-glycoprotein (Pgp)

Expression of Pgp could affect the functions of several transporters. Previous studies have shown a specific relationship between Pgp and swelling-activated Cl^- currents (_Cls), and suggested that Pgp is a Cl^- channel regulator (EMBO J., 14:68-75; Am. J. Physiol., 270:C1370-C1378). Activation of protein kinase C (PKC) inhibits I_Cl^s only in Pgp-expressing cells. In these cells, Pgp expression was obtained by transfection and subsequent selection with chemotherapeutic agents or using a vaccinia virus expression system. Here, we report that activation of PKC with phorbol 12-myristate 13-acetate (PMA) inhibits I_Cl^s in cells transfected with Pgp cDNA, and selected without exposure to chemotherapeutic agents. In Pgp(+) cells, PMA (200 nM, 5 min prior to cell patch-clamp) reduced I_Cl^s from 208 ± 31 to 107 ± 28 pA/pF (all I_Cl^s values listed were measured at + 80 mV). Exposure of the Pgp(-) parental cells to PMA did not alter I_Cl^s. In a Pgp mutant in which serines 661, 667 and 671 were substituted by alanines, PMA had no effect: 262 ± 52 without PMA, and 260 ± 26 pA/pF with PMA. In Pgp(+) cells, 8-Br-cAMP reduced I_Cl^s from 254 ± 22 to 109 ± 3 pA/pF. These results, in conjunction with observations in cells expressing CFTR and the sulfonylurea receptor, suggest that phosphorylation of ABC proteins may be a widespread mechanism of regulation of membrane transporters.